Search PubMed⌕ Search

Biomedical subjects

S K Dower

Publications and source records attributed to S K Dower.

At least 127 records · Page 7Linked to original sources
PubMed1984

The interaction of monoclonal antibodies with MHC class I antigens on mouse spleen cells. I. Analysis of the mechanism of binding.

We studied the mechanism of binding of radiolabeled, monoclonal anti-H-2 antibodies to mouse spleen cells to determine the number of H-2 antigen molecules per cell. Equilibrium and kinetic data were analyzed in detail according to theoretical models developed for different modes of antibody binding. The results of binding experiments from three monoclonal IgG antibodies (36-7-5, anti-Kk; 27-11-13, anti-DbDd; and 11-4-1, anti-Kk) and their F(ab')2 and F(ab') fragments show that for the IgG and F(ab')2 from all three antibodies, the monovalently and bivalently bound states of the antibody co-exist in rapid equilibrium with one another on the cell surface, with the bivalent state predominating. We show that the relative proportions of the monovalently and bivalently bound species can be estimated from dissociation kinetics experiments, and that once the mode of antibody binding has been established, the density of H-2 determinants on the cell surface can be estimated from equilibrium-binding data. We conclude that the average numbers of H-2K and H-2D molecules on B10.A spleen cells are 5 X 10(4) and 1.1 X 10(5) molecules/cell, respectively.

Animals↗
PubMed1983

The role of non-immune IgG in controlling IgG-mediated effector functions.

The majority of evidence supports the conclusion that IgG-dependent effectors respond to antibodies which have been polymerized artificially or by polyvalent antigens, but not to monomeric IgG antibodies. Effectors can distinguish polymerized IgG antibodies from monomeric IgG because they contain multiple receptor units and can interact multivalently with polymerized IgG. However, monomeric IgG is present at very high concns in plasma and interstitial fluids and will inhibit multivalent interactions in vivo between polymerized antibody and effectors. Such inhibition raises the question of how IgG-mediated effector responses could function in vivo. In this review we present a mathematical model which quantitatively predicts how polyvalent ligands interact multivalently with receptors in the presence of excess monovalent ligand. We then show that results from experiments in vitro using such diverse systems as the binding and endocytosis of immune complexes by macrophages, complement-mediated lysis of antibody-coated target cells, and ADCC can be explained qualitatively by the model. We conclude that monomeric IgG does not totally inhibit IgG-mediated effector functions but, rather, raises the threshold of antibody binding which is required to elicit a response. We then consider how non-immune IgG may serve as a homeostatic regulator of IgG-dependent responses, in vivo, perhaps for the purpose of inhibiting responses to low levels of cell-bound IgG autoantibodies.

Adult↗
PubMed1982

Monoclonal anitbodies in the lymphatics: toward the diagnosis and therapy of tumor metastases.

Monoclonal antibodies subcutaneously injected into mice track to regional lymph nodes and specifically label target cells there. The lymphatic route of administration can be expected to provide much higher sensitivity, higher target-to-background ratio, faster localization, and lower toxicity than the intravenous route when the aim is to diagnose or treat tumor metastases or lymphoma in the lymph nodes.

Animals↗
PubMed1981

Mechanism of binding of multivalent immune complexes to Fc receptors. 2. Kinetics of binding.

The kinetics of association and dissociation of affinity cross-linked IgG oligomers with Fc receptors on P388D1 cells have been studied at 0 degrees C. For dimers prepared with two different affinity cross-linking reagents (DIBADL and BDPE) and also trimers and a heavy oligomer fraction prepared with BDPE, monomeric IgG accelerates the rate of dissociation of bound oligomer from the cells. This observation is consistent with a binding mechanism in which monovalently, and multivalently bound species are in rapid equilibrium with one another on the cell surface and in which the rate of formation of the divalently bound species is faster than the dissociation of the monovalent species from the cells. As predicted, the rates of dissociation of BDPE cross-linked oligomers decrease with size, and dimers (cross-linked with DIBADL) which can self-aggregate on the cell surface dissociate more slowly than nonaggregating (BDPE) dimers. The association and dissociation of oligomers with P388D1 cells proceed by parallel fast and slow processes, even with cells that bind dimers with a single affinity. The origin of this heterogeneity is not known but could arise from a fraction of receptors in environments with limited accessibility to bulk solution. The fast component of the association reaction is second order, and the rate-limiting step of this process is the formation of the monovalently bound intermediate from solution-phase oligomer.

Animals↗
PubMed1981

Small-angle neutron scattering studies of the conformation of myeloma protein MOPC315 and its Fab fragment, and the interaction with a monovalent dinitrophenyl hapten.

The first small-angle scattering study of an immunoglobulin A is reported. Neutron measurements have been made to determine conformational parameters of the mouse myeloma protein MOPC315 and to relate these to previous immunoglobulin G results. Use of the contrast method shows that the MOPC315 IgA molecule is not simply globular, that it has a dry volume of 220.0 +/- 4.5 nm3 corresponding to a mass density of 1.275 +/- 0.025 g cm-3 and that its full and cross-sectional radii of gyration, corrected for concentration dependence, are 7.97 +/- 0.07 nm, 2.40 +/- 0.08 nm and 1.33 +/- 0.07 nm respectively. Similar study of its Fab fragment gives a dry molecular volume of 69.0 +/- 0.7 nm3, a mass density of 1.285 +/- 0.015 g cm-3 and uncorrected radii of gyration that are consistent with those of the parent and support an overall "T" or "Y" conformation in solution. Addition to saturation of a small monovalent dinitrophenyl hapten leaves the dry volume of the whole molecule unaltered, but may slightly lower one or more of its radii of gyration. The significance of this finding is discussed. Comparative studies with rabbit anti-dinitrophenyl immunoglobulin G antibody suggest a different initial conformation but similar consequences of hapten binding, which, if real, are probably unrelated to classical complement fixation.

Amino Acid Sequence↗
PubMed1980

The role of antibody multivalency in immune effector processes.

The preparation of covalently crosslinked oligomers of antibody molecules is described. It is shown that these oligomers can be used to study quantitatively the effects of multivalency on binding to cell surface Fc receptors. It is also shown that three effector systems differ in the ways in which they interact with multivalent immune complexes.

Allosteric Regulation↗
PubMed1979

Crystallization of the Fv fragment of mouse myeloma protein M315.

The Fv fragment of mouse myeloma protein M313 was crystallized from poly(ethylene glycol) solution in the form of monoclinic crystals, space group C2 and unit cell dimensions a = 5.96 nm (59.6 A), b = 5.66 nm (56.6 A), c = 13.79 nm (13.9 A) and beta = 99.7 degrees. Some unusual effects of poly(ethylene glycol)on protein crystals were noted and are discussed.

Animals↗
PubMed1978

The binding of 2,4,6-trinitrophenyl derivatives to the mouse myeloma immunoglobulin A protein MOPC 315.

The binding of Tnp (2,4,6-trinitrophenyl) derivatives to the Fv fragment (variable region of heavy and light chains) of the mouse myeloma IgA protein MOPC 315 was investigated by 270MHz proton nuclear magnetic resonance. Two of the haptens, Tnp-glycine and Tnp-l-aspartate, are in fast exchange with the Fv fragment, and the changes in chemical shifts for both protein and hapten resonances were determined by titrations. For the tightly binding hapten epsilon-N-Tnp-alpha-N-acetyl-l-lysine, which is in slow exchange with the Fv fragment, the changes in chemical shifts for the hapten H(3)+H(5) resonances were determined by cross-saturation. By using these data and the known structure of the combining site of protein MOPC 315 [Dwek, Wain-Hobson, Dower, Gettins, Sutton, Perkins & Givol (1977), Nature (London) 266, 31-37] the mode of binding of Tnp derivatives is deduced by ring-current calculations. The trinitrophenyl ring stacks with tryptophan-93(L) (light chain) in the ;aromatic box' formed by tryptophan-93(L), tyrosine-34(L) and phenyl-alanine-34(H) (heavy chain). Further evidence for the stacking interaction with a tryptophan residue is provided by the similarity of the optical-difference spectra observed with Tnp-aminomethylphosphonate in the presence of either the Fab fragment (light chain and N-terminal half of heavy chain) of protein MOPC 315 or tryptophan. These data show that the modes of binding of all the Tnp derivatives are very similar, despite a 100-fold range in their affinities. It is also concluded that the modes of binding of Dnp (2,4-dinitrophenyl) and Tnp derivatives to protein MOPC 315 are very similar, and that the structural basis for this is that the aromatic box is large enought to allow the trinitrophenyl ring to stack with tryptophan-93(L) while still forming hydrogen bonds to asparagine-36(L) and tyrosine-34(L).

Animals↗
PubMed1977

The combining site of the dinitrophenyl-binding immunoglobulin A myeloma protein MOPC 315.

Magnetic-resonance techniques are used to refine the model of the combining site of the Fv fragment of the dinitrophenyl-binding mouse myeloma protein MOPC 315 constructed by Padlan, Davies, Pecht, Givol & Wright (1976) (Cold Spring Harbor Symp. Quant. Biol.41, in the press). Light-absorption studies indicate a dinitrophenyl-tryptophan interaction in the Fv fragment of the type occurring in free solution. The Dnp-aspartate-tryptophan complex is therefore used as a starting point for the n.m.r. (nuclear-magnetic-resonance) analysis of the dinitrophenyl-Fv fragment interaction. Ring-current calculations are used to determine the geometry of the complex. The specificity of complex-formation between dinitrophenyl and tryptophan is confirmed by the lack of ring-current shifts of the dinitrophenyl resonances when tryptophan is replaced by any other aromatic amino acid. Proton n.m.r. difference spectra (at 270MHz), resulting from the addition of a variety of haptens to the Fv fragment, show that the combining site is highly aromatic in nature. Calculations on the basis of ring-current shifts define the geometry of the combining site, which involves a dinitrophenyl ring in van der Waals contact with four aromatic amino acid residues on the protein. The observation of a nuclear Overhauser effect on the H((3)) resonance of the dinitrophenyl ring provides additional constraints on the relative geometry of the H((3)) proton and an aromatic amino acid residue on the Fv fragment. The specificity of the Fv fragment for dinitrophenyl ligands arises from a stacking interaction of the dinitrophenyl ring with tryptophan-93(L), in an ;aromatic box' of essentially tryptophan-93(L), phenylalanine-34(H) and tyrosine-34(L); asparagine-36(L) and tyrosine-34(L) also contribute by forming hydrogen bonds with the nitro groups on the dinitrophenyl ring. The n.m.r. results also confirm that the antibody-hapten reaction may be visualized as a single encounter step. An Appendix shows the method of calculation of ring currents for the four aromatic amino acids and their use in calculating structures.

Binding Sites, Antibody↗
PubMed1977

Specificity of interactions of hapten side chains with the combining site of the myeloma protein MOPC 315.

The pKa values of the three histidine residues in the Fv fragment (variable region of the heavy and light chains) of the mouse myeloma protein MOPC 315, measured by high resolution n.m.r. (nuclear magnetic resonance), are 5.9, 6.9 and 8.2. The perturbation of the pKa of one of the histidines (pKa 6.9) on the addition of hapten and the narrow linewidth of its proton resonances suggests that it is at the edge of the combining site. References to the model of the Fv fragment [Padlan, Davies, Pecht, Givol & Wright (1976) Cold Spring Harbor Symp. Quant. Biol. 41, in the press] allows assignment of the three histidine residues, histidine-102H, histidine-97L and histidine-44L. The determination of the pKa of the phosphorus group, by 31P n.m.r., of a homologous series of Dnp- and Tnp- (di- and tri-nitrophenyl) haptens has located a positively charged residue. Molecular-model studies on the conformations of these haptens show that the residue is at the edge of the site. The model suggests that the positively charged residue is either arginine-95L or lysine-52H.

Antibody Specificity↗
PubMed1975

The binding of lanthanides to non-immune rabbit immunoglobulin G and its fragments.

The binding of Gd(III) to rabbit IgG (immunoglobulin G) and the Fab (N-terminal half of heavy and light chain), (Bab')2 (N-terminal half of heavy and light chains joined by inter-chain disulphide bond), Fc (C-terminal half of heavy-chain dimer)and pFc' (C-terminal quarter of heavy-chain dimer) fragments was demonstrated by measurements of the enhancement of the solvent-water proton relaxation rates in the appropriate Gd(III) solutions. At pH 5.5 there are six specific Gd(III)-binding sites on the IgG. These six sites can be divided into two classes; two very 'tight' sites on the Fc fragment (Kd approx. 5 muM) and two weaker sites on each Fab region (Kd approx. 140 muM). Ca(II) does not apparently compete for these metal-binding sites. The metal-binding parameters for IgG can be explained as the sum of the metal binding to the isolated Fab and Fc fragments, suggesting that there is no apparent interaction between the Fab and Fc regions in the IgG molecule. The binding of Gd(III) to Fab and Fc fragments was also monitored by measuring changes in the electron-spin-resonance spectrum of Gd(III) in the presence of each fragment and also by monitoring the effects of Gd(III) on the protein fluorescence at 340 nm (excitation 295 nm). The fluorescence of Tb(III) solutions of 545 nm (excitation 295 nm) is enhanced slightly on addition of Fab or Fc.

Animals↗
PubMedDate not supplied

The cell surface receptors for interleukin-1 alpha and interleukin-1 beta are identical.

Interleukin-1 (IL-1) is a factor that can induce proliferation of murine T lymphocytes and can elicit a variety of other biological responses. These include bone resorption, fibroblast proliferation, acute phase protein release from hepatocytes, cartilage breakdown and fever. This spectrum of activities is consistent with a role for IL-1 as a mediator of inflammation. Recently, sequence data have shown that there are at least two members of the IL-1 family; these distantly related proteins have been termed IL-1 alpha and IL-1 beta. We have found previously that both murine T cells and fibroblasts possess a relative molecular mass (Mr) approximately 80,000 (80K) plasma membrane receptor for human IL-1 beta. We show here that the receptor for IL-1 alpha on both murine and human cells is identical to that for IL-1 beta. This result raises the issue of what separation, if any, there might be between the biological activities of IL-1 alpha and IL-1 beta.

Animals↗